铁螯合酶的研究。硫醇及其他因素对活性测定的影响。
Studies on ferrochelatase. The effects of thiols and other factors on the determination of activity.
作者信息
Porra R J, Vitols K S, Labbe R F, Newton N A
出版信息
Biochem J. 1967 Aug;104(2):321-7. doi: 10.1042/bj1040321.
Abstract
- Haems are unstable under aerobic conditions in the presence of thiols, which are used to activate the ferrochelatase enzyme; catalase inhibits this degradation of haem. In addition, thiols interfere with the determination of protohaem as its pyridine haemochromogen derivative. 2. Three ferrochelatase assays are described that minimize interference by these two reactions. Two of these assays involve measurement of porphyrin utilization, one spectrophotometrically and the second spectrofluorimetrically. The third assay measures haem formation by a pyridine haemochromogen technique. Results obtained with these three methods were in close agreement at a GSH concentration of 4mm. 3. The stimulatory effect of GSH on ferrochelatase has been confirmed. The spectrum of the haem formed is dependent on GSH concentration; at high GSH concentrations (20mm) the haem is in the reduced state, but at low concentration (4mm) the spectrum of the product resembles that of an oxidized haemoprotein such as ferrihaemoglobin. 4. The inhibitory effect of oxygen on ferrochelatase activity has been confirmed by spectrophotometric assay of porphyrin disappearance.
摘要
- 在有氧条件下，血红素在硫醇存在时不稳定，硫醇用于激活亚铁螯合酶；过氧化氢酶可抑制血红素的这种降解。此外，硫醇会干扰原血红素作为其吡啶血色原衍生物的测定。2. 描述了三种亚铁螯合酶测定方法，可将这两种反应的干扰降至最低。其中两种测定方法涉及卟啉利用的测量，一种是分光光度法，另一种是荧光分光光度法。第三种测定方法通过吡啶血色原技术测量血红素的形成。在谷胱甘肽浓度为4mM时，用这三种方法获得的结果非常一致。3. 谷胱甘肽对亚铁螯合酶的刺激作用已得到证实。形成的血红素光谱取决于谷胱甘肽浓度；在高谷胱甘肽浓度（20mM）下，血红素处于还原状态，但在低浓度（4mM）下，产物的光谱类似于氧化血红蛋白（如高铁血红蛋白）的光谱。4. 通过分光光度法测定卟啉消失，证实了氧气对亚铁螯合酶活性的抑制作用。
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